Imperial College Centre for Synthetic Biology, London, SW7 2AZ, U.K.
Department of Chemistry, Imperial College London, London, SW7 2AZ, U.K.
ACS Synth Biol. 2022 Feb 18;11(2):562-569. doi: 10.1021/acssynbio.1c00580. Epub 2022 Feb 8.
Synthetic RNA systems offer unique advantages such as faster response, increased specificity, and programmability compared to conventional protein-based networks. Here, we demonstrate an RNA-based toggle switch using RNA aptamers capable of inhibiting the transcriptional activity of T7 or SP6 RNA polymerases. The activities of both polymerases are monitored simultaneously by using Broccoli and malachite green light-up aptamer systems. In our toggle switch, a T7 promoter drives the expression of SP6 inhibitory aptamers, and an SP6 promoter expresses T7 inhibitory aptamers. We show that the two distinct states originating from the mutual inhibition of aptamers can be toggled by adding DNA sequences to sequester the RNA inhibitory aptamers. Finally, we assessed our RNA-based toggle switch in degrading conditions by introducing controlled degradation of RNAs using a mix of RNases. Our results demonstrate that the RNA-based toggle switch could be used as a control element for nucleic acid networks in synthetic biology applications.
合成 RNA 系统相较于传统的基于蛋白质的网络具有更快的反应速度、更高的特异性和可编程性等独特优势。在这里,我们展示了一种基于 RNA 的 toggle switch,它使用能够抑制 T7 或 SP6 RNA 聚合酶转录活性的 RNA 适体。通过使用 Broccoli 和孔雀石绿点亮适体系统,同时监测两种聚合酶的活性。在我们的 toggle switch 中,T7 启动子驱动 SP6 抑制适体的表达,而 SP6 启动子表达 T7 抑制适体。我们表明,来自适体相互抑制的两种不同状态可以通过添加 DNA 序列来封闭 RNA 抑制适体来切换。最后,我们通过使用混合核糖核酸酶引入 RNA 的受控降解,在降解条件下评估了我们的基于 RNA 的 toggle switch。我们的结果表明,基于 RNA 的 toggle switch 可用作合成生物学应用中核酸网络的控制元件。